The consequences of specific immunodeficiency, the lack of a specific immune response to a pathogen, results in significant mortality from bacterial infection in transplant recipients with chronic graft-versus-host disease (GVHD). Immunodeficiency in these patients cannot be attributed to low numbers of B cells or low levels of serum IgM and IgG, suggesting that the specificity of response or "antibody repertoire" may be limited. The goal of this research project is to determine whether a restricted antibody repertoire contributes to humoral immunodeficiency following bone marrow transplantation (BMT). We have shown in preliminary studies some nonrandom usage of immunoglobulin gene segments in BMT recipients. The specific gaol of this project are to 1) dissect the evolution of the antibody repertoire in B cells from recipients with and without chronic GVHD; and 2) molecularly evaluate the antibody responses of recipients receiving various Hemophilus influenzae vaccines (project 0011). In aim 1, we will isolate B cells, purify DNA, and amplify immunoglobulin gene rearrangements by polymerase chain reaction. The usage of different variable genes will be evaluated by hybridization with VH family-specific probes and determining the DNA sequence. In aim 2, B cell lines synthesizing anti-Hemophilus influenzae antibodies will be established. DNA sequence analysis will be performed on both the heavy and light chains of these cell lines. The usage of immunoglobulin genes in normal individuals and BMT recipients following T cell-independent and T cell-dependent vaccines will be determined. We will test whether a repertoire deficit is responsible for nonresponsiveness to vaccines. These studies will directly assess the role of the utilized immunoglobulin repertoire in specific humoral immunodeficiency. Results of these experiments will enable a deeper evaluation of present therapy and contribute to the design of new therapies.